Detailed proteomic analysis of biomolecule samples such as serum or cell extract could provide valuable information regarding the person's health or biological cells under study. However, current sample preparation technologies cannot deal with the diversity and dynamic range of these biomolecule samples effectively, and therefore the detection of information-rich minority protein species (signaling molecules and cytokines) is severely limited. Efficient protein separation in microfluidic systems could provide a solution for this challenge, but the limitation in sample volume, as well as the lack of molecular signal amplification strategy, has been major issues. Recent advances in nanofluidics provide useful tools for proteomic sample preparation, to address these challenges. Nanofluidic filters could be used for a gel-free separation of proteins based on the size. Permselective ion current generated by the nanofluidic channel was used to concentrate proteins and peptides from a dilute solution, to increase the sample volume capacity of the microfluidic sample preparation system. Also, continuous-flow pI-based biomolecule sorting system was demonstrated. In this project, we will develop these nanofluidic components further, and integrate them into an automatic proteomic sample preparation system that could sort out complex biomolecule sample based on the property of the target molecule. The flexibility and robustness of the micro/nanofluidic sample preparation devices that will be developed from this project will facilitate the system integration greatly. The resulting sample preparation microdevices could be easily integrated with various types of detectors, including antibody-based biosensor or mass spectrometry. Such an integration will enhance the selectivity and sensitivity of the biodetection by orders of magnitudes, therefore impacting various fields such as human health monitoring, biomarker detection, and systems biology.